Even with increased restore Virus persists in tissues with the potential to become reactivated when ART is discontinued underscoring a major challenge of HIV eradication. The possibility of curative treatment for HIV-1 infection has been energized by the ?Berlin patient?, who received stem cell transplants from a CCR5?32-homozygous donor after radiation and chemotherapy for his acute myelocytic leukemia and has been undetectable for the virus in his blood and tissues for at least 10 years since stopping antiretroviral therapy (ART) after the transplants. In our previous studies, we successfully applied CRISPR/Cas9 gene editing to target and completely eradicate integrated HIV sequences in in vitro cell culture models, ex vivo cultured HIV+ patient derived cells and in vivo using HIV transgenic mice and rats and most recently in close collaboration with Dr. Howard Gendelman's team (UNMC) in ART treated HIV-infected humanized mice. In up to a third of infected treated humanized mice, virus was not detected in blood, spleen, lung, kidney, liver, gut- associated lymphoid tissue and brain by ultrasensitive nested and digital droplet PCR and RNAscope tests. No viral rebound was demonstrated after ART cessation. In this application, we will employ a gene editing strategy for targeting SIV and CCR5 in SIV-infected non-human primate model. In light of our preliminary data and published studies, our central hypothesis is that employment of a combined CRISPR/Cas9 gene editing platform can effectively excise SIV proviral DNA from the latent viral reservoir and by editing CCR5 gene prevent spread of the virus and reinfection in the animal leading to a reduction in the functional viral reservoir contemporary anti-retroviral therapy (ART) regimens the rate of survival, HIV remain an enormous health immune health and is not curative. that suppress viral replication burden . Unfortunately, ART does and have not fully and/or sterile cure. To in in vivo SIV animal eradicating virus from host genomes inducible viral RNA reservoir In this application, we will establish SIV and CCR5 gene editing platforms in SIV-infected non-human primates (Aim 1) and determine the immunological and virological effects and the underlying mechanism of cure or delayed viral rebound after gene editing and antiretroviral therapy interruption (ATI) (Aim 2). We will use single and then a sequential dual editing that targets the cellular gene (CCR5) followed by the proviral DNA (SIV). Studies in this application will this end, a team of experienced investigators has been assembled with unique expertise models and immunology (T. Burdo, Temple University), (K. Khalili, Temple University), and (J. Karn, Case Western Reserve University). gene editing technologies for state of the art assays measuring the allow us to gain valuable insight into the ability of CRISPR/Cas9 gene editing for cure strategies. These studies infection, in vivo are highly translatable and will provide knowledge toward the eradication or functional cure of HIV off ART without detectable plasma viremia orthe ultimate goal being to achieve a prolonged period otherevidence of active infection.